Acute pulmonary dysfunction post-bone marrow transplant (BMT) is a relatively frequent and severe complication of allogeneic BMT occurring in the first 100 days with an incidence of 35% in unrelated donor (URD) and 22% in peripheral blood stem cell transplants. Idiopathic Pneumonia Syndrome (IPS) injury represents a subset of these patients that have diagnostic criteria including signs and symptoms of pneumonia, evidence for nonlobar radiographic infiltrates, abnormal pulmonary function, and absence of lower respiratory tract infection. Conditioning regimen injury is the highest contributory factor for IPS. A multivariant analysis of the Seattle data indicated a hazard ratio of 9.3 for IPS in patients >age 40 who received 12 Gray total body irradiation (TBI) as compared to those receiving non-myeloablative transplants. Although acute graft versus host disease (GVHD) is a risk factor, IPS cannot be attributable simply to an alloresponse in humans. The incidence of IPS in allotransplants ranges from 5-20%. A recent large retrospective analysis indicated a rate of 8.4% in conventional transplant recipients. In most studies, the incidence of IPS in humans has been shown to be higher in recipients given more intense conditioning regimens and in recipients of allogeneic vs autologous BMT. Because of the association of alloresponses with IPS injury, the incidence of IPS will probably increase as the donor pool is extended to include BMT from unrelated donors. Once IPS has developed, the death rate is very high (usually ≧75%) and time to mortality is rapid (usually 2 weeks).
Therapeutic strategies are limited and in general consist of supportive care, mechanical ventilation and high dose steroids, which are not sufficient to prevent the high mortality rate. Acute IPS injury also may set the stage for bronchiolitis obliterans (BO) or bronchiolitis obliterans with organizing pneumonia (BOOP) which occur in 5-25% of long-term allo-BMT survivors and has been linked to chronic GVHD. Some cases of IPS, BO and BOOP will result in fibrosis which may occur within several weeks after onset. While tumor necrosis factor (TNF) neutralization has had some success in IPS management, a need for more effective clinical treatment exists.
Stem Cells
The quintessential stem cell is the embryonal stem (ES) cell, as it has unlimited self-renewal and can differentiate into all tissue types. ES cells are derived from the inner cell mass of the blastocyst, or primordial germ cells from a post-implantation embryo (embryonal germ cells or EG cells). ES and EG cells have been derived from mouse, and more recently also from non-human primates and humans. When introduced into mouse blastocysts or blastocysts of other animals, ES cells can contribute to all tissues of the mouse (animal). When transplanted in post-natal animals, ES and EG cells generate teratomas.
ES (and EG) cells can be identified by positive staining with antibodies to SSEA 1 (mouse) and SSEA4 (human). At the molecular level, ES and EG cells express a number of transcription factors specific for these undifferentiated cells. These include oct-4 and rex-1. Also found are the LIF-R and the transcription factors sox-2 and rox-1. Rox-1 and sox-2 are also expressed in non-ES cells. Another hallmark of ES cells is the presence of telomerase, which provides these cells with an unlimited self-renewal potential in vitro.
Oct-4 is a transcription factor expressed in the pregastrulation embryo, early cleavage stage embryo, cells of the inner cell mass of the blastocyst, and in embryonic carcinoma (EC) cells (Nichols J., et al (1998) Cell 95:379-91). Oct-4 is down-regulated when cells are induced to differentiate in vitro. In the adult animal oct-4 is only found in germ cells. Several studies have shown that oct-4 is required for maintaining the undifferentiated phenotype of ES cells and plays a major role in determining early steps in embryogenesis and differentiation. Oct-4, in combination with Rox-1, causes transcriptional activation of the Zn-finger protein rex-1, which is also required for maintaining ES in an undifferentiated state ((Rosfjord E, Rizzino A. (1997) Biochem Biophys Res Commun 203:1795-802; Ben-Shushan E, et al (1998) Mol Cell Biol 18:1866-78)). Likewise, sox-2, is needed together with oct-4 to retain the undifferentiated state of ES/EC (Uwanogho D et al (1995) Mech Dev 49:23-36) and to maintain murine (but not human) ES cells. Human or murine primordial germ cells require the presence of LIF.
The oct 4 gene (oct 3 in humans) is transcribed into at least two splice variants in humans, oct3A and oct3B. The oct3B splice variant is found in many differentiated cells whereas the oct3A splice variant (also previously designated oct3/4) is reported to be specific for the undifferentiated embryonic stem cell. See Shimozaki et al. Development 130:2505-12 (2003).
Adult stem cells have been identified in most tissues. Hematopoietic stem cells are mesoderm-derived and have been purified based on cell surface markers and functional characteristics. The hematopoietic stem cell, isolated from bone marrow, blood, cord blood, fetal liver and yolk sac, is the progenitor cell that reinitiates hematopoiesis for the life of a recipient and generates multiple hematopoietic lineages. Hematopoietic stem cells can repopulate the erythroid, neutrophil-macrophage, megakaryocyte and lymphoid hemopoietic cell pool. Stem cells which differentiate only to form cells of hematopoietic lineage, however, are unable to provide a source of cells for repair of other damaged tissues, for example, heart.
Neural stem cells were initially identified in the subventricular zone and the olfactory bulb of fetal brain. Several studies in rodents, and more recently also non-human primates and humans, have shown that stem cells continue to be present in adult brain. These stem cells can proliferate in vivo and continuously regenerate at least some neuronal cells in vivo. When cultured ex vivo, neural stem cells can be induced to proliferate, as well as to differentiate into different types of neurons and glial cells. When transplanted into the brain, neural stem cells can engraft and generate neural cells and glial cells.
Mesenchymal stem cells, originally derived from the embryonal mesoderm and isolated from adult bone marrow, can differentiate to form muscle, bone, cartilage, fat, marrow stroma, and tendon. During embryogenesis, the mesoderm develops into limb-bud mesoderm, tissue that generates bone, cartilage, fat, skeletal muscle and possibly endothelium. Mesoderm also differentiates to visceral mesoderm, which can give rise to cardiac muscle, smooth muscle, or blood islands consisting of endothelium and hematopoietic progenitor cells. Primitive mesodermal or mesenchymal stem cells, therefore, could provide a source for a number of cell and tissue types. Of the many mesenchymal stem cells that have been described, all have demonstrated limited differentiation to form only those differentiated cells generally considered to be of mesenchymal origin. To date, the best characterized mesenchymal stem cell reported is the cell isolated by Pittenger, et al. (1999) and U.S. Pat. No. 5,827,740 (SH2+ SH4+ CD29+ CD44+ CD71+ CD90+ CD106+ CD120a+ CD124+ CD14− CD34− CD45−). This cell is capable of differentiating to form a number of cell types of mesenchymal origin, but is apparently limited in differentiation potential to cells of the mesenchymal lineage, as the team who isolated it noted that hematopoietic cells were never identified in the expanded cultures.